Detection of DNA Damage Caused by Cryopreservation using a Modified SCGE in Large Yellow Croaker, Pseudosciaena crocea
We used single-cell gel electrophoresis (SCGE) to detect the integrity of sperm DNA of the teleost large yellow croaker, Pseudosciaena crocea, cryopreserved with Cortland solution and a range of 5% to 30% DMSO concentrations in order to test how sperm cryopreservation affected the DNA stability of nuclei. Electrophoresis was conducted for 60 min at 130 mA and 15 V. The comet images were analyzed with software CometScore 1.5, and parameters such as comet length, tail length and percentage DNA in the tail were obtained. Then the comet rate and damage coefficient were calculated. Results demonstrated that there were no significant differences in motility, comet rate and damage coefficient between fresh sperm and cryopreserved sperm stored in 5%, 10%, 15% and 20% DMSO, while the sperm cryopreserved with 25% and 30% DMSO had a lower motility, higher comet length and damage coefficients than those of fresh sperm. There was a positive correlation between comet rate of cryopreserved sperm and the concentration of DMSO. Our results demonstrate that toxicity of the cryoprotectant is the main cause of DNA damage in cryopreserved sperm nuclei.
KeywordsPseudosciaena crocea sperm cryopreservation genetic damage single-cell gel electrophoresis
We thank the following projects for their support: K. C. Wong Magna Fund in Ningbo University, The Scientific and Technical Project of Zhejiang Province, National Natural Science Foundation of China (Grant Nos. 41276151, 31272642 and 31072198).
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